The bacterial genus Mycobacterium, which consists of over 120 known species, is a major burden of infectious disease in humans. As the most pathogenic species, Mycobacterium Tuberculosis Complex (MTBC), including Mycobacterium tuberculosis, Mycobacterium africanum, Mycobacterium microti and Mycobacterium bovis, infects around one-third of the world's population and claims over 1.5 million lives each year (Dye et al., (2008) Lancet Infect. Dis. 8: 233-243; Zumla et al., (2013) N. Engl. J. Med. 368: 745-755; Anonymous (2013) Global Tuberculosis Report 2013, World Health Organization).
Tuberculosis is a highly infectious airborne disease. A rapid and accurate detection of MTBC is considered to be the most critical step in containing the spread and decreasing the mortality rate of this disease (Urdea et al., (2006) Nature 444 (Suppl. 1): 73-79; Keeler et al., (2006) Nature: 49-57; McNerney & Daley (2011) Nat. Rev. Microbiol. 9: 204-213; Dheda et al., (2013) Respirology 18: 217-232). Other than MTBC, Non-Tuberculosis Mycobacteria (NTM), such as slow-growing Mycobacterium avium and Mycobacterium intracellulare, can also cause skin disease, Johne's disease, inflammatory bowel disease, and Crohn's disease with relatively high morbidity and mortality.
The slow growth rate of Mycobacterium tuberculosis presents a large hurdle that needs to be overcome for the effective detection of these virulent pathogens. So far, no diagnostic assay is cost-effective and can also provide results within a single patient-health-care visit (within a couple of hours) (Keeler et al., (2006) Nature: 49-57; McNerney & Daley (2011) Nat. Rev. Microbiol. 9: 204-213; Dheda et al., (2013) Respirology 18: 217-232). Current diagnosis such as the gold standard culture-based technique, smear microscopy, and nucleic acid-based diagnostic methods are time-consuming, expensive, or technically demanding.
The intrinsic resistance of mycobacteria to β-lactam antibiotics worsens the global crisis and treatment options. Among all the resistance mechanisms, β-lactamase is believed to be the key contributor. Since the discovery of the first β-lactamase in 1940, a large number of β-lactamases have been identified that can hydrolyze a variety of β-lactam antibiotics, from penicillin to cephalosporins and carbapenems (Bush & Jacoby (2010) Antimicrob. Agents Chemother. 54: 969-976).
To assay the activity of β-lactamases, a series of non-specific fluorogenic and luminogenic probes has been developed that take advantage of the high sensitivity of fluorescence and luminescence detecting mechanism (Zlokarnik et al., (1998) Science 279: 84-88; Gao et al., (2003) J. Am. Chem. Soc. 125: 11146-11147; Xing et al., (2005) J. Am. Chem. Soc. 127: 4158-4159; Yao et al., (2007) Angew. Chem. Int. Edit. 46: 7031-7034; Kong et al., (2010) Proc. Natl. Acad. Sci. U.S.A. 107: 12239-12244; Rukavishnikov et al., (2011) Anal. Biochem. 419: 9-16; Zhang et al., (2012) Angew. Chem. Int. Edit. 51: 1865-1868).
Fluorogenic probes for the specific pathogen detection have been developed. For example, by engineering the stereochemistry of cephalosporin a series of fluorogenic probes was developed that showed specificity for Carbapenem-Resistant Enterobacteriaceae (CRE) (Shi et al., (2014) Angew. Chem. Int. Edit. 53: 8113-8116). By targeting BlaC, an ambler class A β-lactamase that is highly conserved in MTBC clinical isolates and plays a key role for the pervasive β-lactam-antibiotic resistance, a series of fluorogenic probes for rapid point-of-care detection of MTBC was developed (Cheng et al., (2014) Angew. Chem. Int. Edit. 53: 9360-9364; Xie et al., (2012) Nat. Chem. 4: 802-809). However, although these probes exhibit advantages as low-cost triage tests for use in the resource-limited areas, they are deficient in the sensitivity and specificity required for use in both microscopy centers and as probes for use with point-of-care methods.